1. Field of the Invention
The present invention relates to a method for disrupting cells and purifying nucleic acid. The present invention further relates to an apparatus for disrupting cells and purifying nucleic acids.
2. Description of the Related Art
Methods of isolating DNA from cells are performed using materials that have a tendency for binding to DNA. For example, materials used for DNA isolation include silica, glass fiber, anion exchange resin and magnetic beads (Rudi, K. et al., Biotechniques 22, 506-511 (1997); and Deggerdal, A. et al., Biotechniques 22, 554-557 (1997)). Furthermore, for the purpose of avoiding manual operation and eliminating operator error, several automatic machines have been developed for high-throughput DNA extraction.
The production of high purity double-stranded plasmid DNA, single-stranded phage DNA, chromosomal DNA, and agarose gel-purified DNA fragments is very important in molecular biology. Ideal methods of purifying DNA should be simple and quick, and include minimal manipulation of samples. The DNA molecules obtained using such methods are ready for direct transformation, restriction enzyme analysis, ligation or sequencing. Such methods are very attractive in the automated production of DNA samples. Automated production of DNA samples is often favored in research and diagnosis labs because such methods avoid manual operation and minimize operator error.
Conventionally, methods of purifying nucleic acids using a solid phase are known. For example, U.S. Pat. No. 5,234,809 discloses a method of purifying nucleic acids using a solid phase, e.g., silica particles, to which nucleic acids are bound. The method of U.S. Pat. No. 5,234,809 includes mixing a starting material, a chaotropic material, and a nucleic acid binding solid phase; separating the solid phase with the nucleic acid bound thereto from the liquid, and washing the solid phase nucleic acid complexes. One disadvantage of this method is that the method is time consuming and complicated, and therefore not suitable for a Lab-On-a-Chip (LOC). Furthermore, another disadvantage of the method is that a chaotropic material is used.
U.S. Pat. No. 6,291,166 discloses a method of archiving nucleic acids using a solid phase matrix. This method is advantageous in that nucleic acids are irreversibly bound to the solid phase matrix. By providing nucleic acids irreversibly bound to the solid phase matrix, it is possible to delay analysis of, or to perform repeated analysis of the nucleic acid solid phase matrix complexes after storage. One disadvantage with this method is that alumina (Al2O3), which has a positively-charged surface, needs to be rendered hydrophilic which requires the use of basic materials, such as NaOH. Another disadvantage with this method is that the nucleic acids are irreversibly bound to the hydrophilic alumina, and thus cannot be separated from the alumina.
U.S. Pat. No. 6,936,414 discloses a method of separating nucleic acid from a test sample. The method comprises contacting a test sample with a metal oxide support material and a binding buffer to form nucleic acid/metal oxide support material complexes; separating the complexes from the test sample; and eluting the nucleic acid from the metal oxide support material. Although the invention disclosed in U.S. Pat. No. 6,936,414 is similar to the present invention in that binding of metal oxide and nucleic acid is used, the invention disclosed in U.S. Pat. No. 6,936,414 differs from the present invention in that the invention disclosed in U.S. Pat. No. 6,936,414 uses a chaotropic salt and a detergent as binding materials when nucleic acids are bound to the metal oxide support. Advantageously, the present invention does not utilize chaotropic salts or detergents.
In an attempt to determine whether a Lab-On-a-Chip integration of cell lysis and nucleic acid purification could be achieved, the inventors studied methods of disrupting cells and purifying nucleic acid based on the conventional techniques described above. The inventors confirmed that a Lab-On-a-Chip integration could be achieved and developed a technique for performing cell enrichment, cell lysis, and a process of purifying nucleic acids in a single chamber.